The objective of this project is to investigate mechanisms by which insulin regulates the growth of rat fetal hepatocytes in primary culture. The overall hypothesis is that insulin action is mediated, at least in part, through effects on the actions of other polypeptide growth factors. Since insulin and growth factor signal transmission both involve reversible phosphorylation of proteins, we will focus on protein phosphorylation and protein phosphatase regulation. The hypotheses to be tested are based in large part on preliminary data which have demonstrated the following: [1] Fetal rat hepatocytes in primary culture grow independently of serum or added mitogens; [2] This factor-beta (TGF-beta); [3] Insulin at physiologic concentrations potentiates serum-independent growth by approximately 50%; [4] This insulin-potentiation is abolished in hepatocytes from growth retarded fetuses. The specific aims of this project include the following: [1] Determine the ontogeny of serum-independent growth of fetal and neonatal rat hepatocytes and the autocrine/paracrine factors responsible for this growth; [2] Investigate mechanisms mediated through effects on insulin-like growth factor (IGF) receptors and binding proteins by which insulin potentiates fetal hepatocyte growth; [3] Investigate mechanisms mediated by protein phosphorylation by which insulin potentiates fetal hepatocyte growth; [4] Study corresponding mechanisms by which glucagon and TGF-beta inhibit fetal hepatocyte growth; [5] Compare the growth regulatory effects and mechanisms of action of insulin, glucagon and TGF-beta in fetal hepatocytes from normal, growth retarded and macrosomic fetuses. We anticipate that these studies will provide insight into the mechanisms by which fetal hepatic growth is augmented in fetuses of diabetic mothers.